Project Summary Research is proposed to address PA-04-119 Biodefense and Emerging Infectious Disease Research Opportunities. WNV is a NIAID Category B priority pathogen. The introduction of a single WNV strain at a relatively well characterized time and place in North America has provided a unique opportunity to prospectively study the evolution of an arthropod-transmitted RNA virus as it adapts to a na[unreadable]ve ecological niche. While WNV in the Americas remains a relatively homogeneous virus population, a single genotype that differs from the introduced genotype has become dominant throughout North America, and since 2002 has displaced previously existing genotypes in the United States. Preliminary studies suggest that the dominance of this new genotype may be facilitated by interactions between the virus and the mosquito host as well as by interactions with natural avian hosts. The range of biological mechanisms that may lead to the displacement of one arboviral genotype with another, however, has not been systematically examined. Accordingly, we will test the hypothesis that dominance of one WNV genotype may result from selective advantages of this genotype in the mosquito and/or avian host. Specifically, proposed studies will focus on whether this dominance is achieved through differential potential for genetically defined WNV strains to (a) be transmitted vertically and subsequently perorally by mosquitoes, (b) infect, produce viremia and viral shedding in ecologically relevant vertebrate hosts. Further, we propose experiments that will determine whether these phenotypic differences are associated with differences at the cellular level in viral replication events. Finally, we will determine the point(s) in the WNV transmission cycle that are likely to have been responsible for the observed extinction of one viral genotype as it has been displaced with another in the U.S. Interactions of an arbovirus with both vertebrate and invertebrate hosts impact the intensity of virus transmission and thereby alter the rate of enzootic amplification, genetic stability of the virus, and subsequent viral fitness. Results from these studies will have direct implications on public health by increasing our understanding of the selective processes that shape invasive vector-borne virus populations using WNV as a model system for a vector-borne disease agent introduced into a naive environment.